denoising vs dereplication

[llenzi (Luca)]

Hi @taf1g17

Both deblur and dada2 should return error-free sequences, which will be beneficial for the taxonomy assignment step. Also a smaller data set to handle after the denoising, because sequences with errors are not discarded but reverted to the original state. You could compensate this via filtering the low abundance OTUs, but you always taking the risk to discard real low abundant clusters as well by applying a filtering by abundance.

There are few interesting discussion on comparing OTUs (clusters obtained by vsearch, as in old qiime1) and ASVs/ESVs (amplicon sequence variants obtained by denoisers as dada2 or deblur), one is:

ESV vs OTU deflation (QIIME1 vs 2)

This is a more general question. Now that we are switching completely to QIIME2, i did one comparison between 97% OTU clustering in QIIME 1 and 99% ESV clustering in QIIME 2/deblur. The dataset is a fairly standard MiSeq 2x250 bp run with >100 bp overlap. I compared two experiments by reads surviving the pipeline raw OTUs/ESVs identified by sample. I found that QIIME2 accepts more reads, while identifying much less ESVs than OTUs. This was surprising because i would expect that 99% clusteri…

Another good one is:

Major differences in alph diversity between q1 and q2

Hi @uria, The short answer: yes, these differences are expected, sensible, and acceptable. @colinbrislawn's answer on the qiime1 forum does a great job of summarizing the cause of this discrepancy — the methods used by that user (and yourself) in qiime1 and qiime2 are very different — but does not go into the nitty gritty details. Let's discuss those here. The long answer: The differences that you are observing are not differences between qiime1 and qiime2, strictly speaking. Each of these i…

Which remind me about chimera filtering steps embedded into deblur and dada2, that seems missing in your pipeline from the step you mention.

One question, what are you going to do next with the dereplicated sequences?

Keep in mind, working with ASVs or OTUs are both accepted way to get your results, so as long you are doing correctly they are both valid! Mostly is a matter of preference and knowing what you are working with!

Luca